The Nerve Growth Factor (NGF) (Levi-Montalcini, 1952) is a pleiotropic neurotrophin having a fundamental role in the neuronal differentiation of central and peripheral nervous system. Accordingly NGF is essential for the differentiation of cholinergic neurons of the basal forebrain, the sensory and sympathetic neurons.
NGF is also necessary in post-differentiation steps, as it is able to modulate cellular apoptosis (Davies, 1992), the synthesis of cytoskeletal elements during neuroaxonal regeneration (Snider, 1989), the synthesis of enzymes, peptides and neurotransmitters (Eide et al., 1993), synaptic rearrangements and neuronal proliferation (Schnell et al., 1994). NGF is thought to play a fundamental role also in neurodegenerative processes and neuronal ageing (Connor & Dragunow, 1998).
The mechanisms regulating such different activities are yet to be clarified: the pleiotropic activity of NGF and the unavailability of adult animal models in which NGF activity is neutralized do not allow to relate its expression to a unique and definite phenotype or prefixed and recognisable function. The inactivation of NGF in adult animal models has been attempted by different approaches, among which immunoneutralisation by systemic delivery of anti-NGF neutralising antibodies (Levi-Montalcini et al., 1960) o gene <<knockout>> in transgenic mice (Crowley et al., 1994).
However in both cases the results were rather disappointing. The systemically delivered antibodies do not cross the blood-brain barrier and the effect of NGF neutralization in the other districts depend on many additional factors which are not easily standardised, like anti-serum titer and affinity, clearance rate or cross-reactivity of polyclonal antibodies with other neurotrophins, etc.
On the other hand the <<knockout>> approach in mice allowed to confirm the importance of said neurotrophin during development and the lack of redundancy of its own intracellular signalling system, but it failed to provide the expected adult model for the study of the NGF: indeed ngf−/− transgenic mice die shortly after birth, before the development of phenotypic changes linked to ageing. The phenotype of ngf+/− heterozygotic mice was also not very instructive, to study the phenotype associated with NGF deficit in adult: the NGF level in the heterozygous mouse is only 25% lower than in the control. This yields only a mild phenotype characterized by a faint cholinergic deficit, without apparent anomalies related to human neurodegenerative pathologies.
However different lines of experimental data suggest that NGF plays a key role in neurodegenerative syndromes (Connor & Dragunow, 1988). Senile dementia and Alzheimer's disease (AD) are neurodegenerative syndromes characterised by a progressive dementia. Alzheimer's disease affects 5% of 70 years old people and more than 30% of 80 years old people. Its incidence, in relation to the improvement of the life conditions and lengthening of the mean age, is destined to double in the next thirty years. Social costs for said pathology are very high. Alzheimer's disease mainly affects neurons of the cerebral cortex and the hippocampus and is characterised both by clinical symptoms (for example, the progressive loss of cognitive capacities) and by characteristic histopathological hallmarks (which can be diagnosed only post-mortem), as the formation of extracellular plaques of β-amyloid protein, neurofibrillar intracellular aggregates (tangles) consisting of hyperphosphorylated tau protein, and <<neuronal loss>> (Goedert, 1993; Mandelkow and Mandelkow, 1993; Selkoe, 1994). In the central nervous system basal forebrain cholinergic neurons are particularly affected, resulting in a decrease of acetylcholine synthesis and selective loss of said neurons. Cholinergic neurons are dependent on NGF, which acts through high (TrkA) and low affinity (p75) receptors.
Furthermore, Alzheimer's disease is associated to motorial disorders due to loss of cognitive capacities and coordination, as well as to pathologies of skeletal muscles such as amyloid deposition in skeletal muscle cells (Fukuchi et al., 1998; Jin et al., 1998). It is interesting to point out that NGF further exerts a function on non-neuronal cells, due to the presence of NGF receptors, namely p75 and TrkA, on non neuronal cells, including muscle cells.
In spite of enormous investments, up to now an early diagnosis and a suitable therapy for AD are unavailable. This is due, above all, to the unavailability of experimental cellular or animal models that mimic in a complete and accurate way the formation of the aberrant neuropathological structures found in AD brains. During recent years different transgenic models have been produced, with the aim of defining the aetiopathogenesis of Alzheimer's disease and of selecting useful compounds for therapy. Based on the histological, immunological, and molecular hallmarks of Alzheimer's disease, such as the presence of deposits of β-amyloid protein in the central nervous system, transgenic animals were obtained wherein the wild-type β-amyloid precursor protein (βAPP) is expressed at levels higher than the endogenous one, or is expressed in a mutated form wherein the mutations are those found in the genetic forms of the disease (Hsiao, WO 97/87492 and Games, WO 96/40896). Other transgenic models relate to animals wherein the transgene is the presenilin-1 or -2 (Citron et al., 1996; Strchler-Pierrat et al., 1997), alone or together with the amyloid precursor protein (APP) (Borchelt et al., 1997; Holcomb et al., 1998; Wong et al., 1999), the tau protein (Gotz et al., 1995; Brion et al., 1999) or a βAPP protein C-99 fragment (Jin et al., 1998).
However so far all models develop only some of the morphological, histological or molecular markers defined as characteristic for the diagnosis of Alzheimer's disease and therefore do not represent complete models, effectively suitable to study and provide successful therapies and test drugs. The lack of a comprehensive model for Alzheimer's disease is recognized to represent a crucial bottleneck limiting the screening and validation of new therapeutical agents.